Cross Reference to Related Applications
This application is related to copending patent application Ser. No. 664,366, filed Mar. 4, 1991, entitled Butadiene Acrylonitrile Polymeric Coating Chromatographic Packing Material, and having common assignee and common inventorship.
1. Field of the Invention
This invention relates to the immobilization and subsequent functionalization of a hydrocarbonaceous polymers on inorganic support materials for subsequent use as chromatographic stationary phases. More specifically, this invention relates to the coating of a polymer onto a metal oxide followed by in situ crosslinking of the polymer and chemical functionalization thereby producing stationary phases that exhibit unique chromatographic selectivities and excellent pH and chemical stability.
2. Prior Art
Chemically modified silica supports are currently the most widely used stationary phases for reversed-phase liquid chromatography. By reversed-phase chromatography it is meant that the adsorbent is less polar than the eluting solvent, and in normal phase chromatography the adsorbent is more polar than the eluting solvent. That is, in reversed-phase chromatography, the more non-polar sample components interact more with the relatively non-polar stationary phase and thus elute later than polar sample components. Typical mobile phases for reversed phased chromatography are aqueous buffers, water, methanol, acetonitrile, tetrahydrofuran, and mixtures of water or buffer with these organic solvents.
However, these chemically modified silica supports, in particular, alkyl-bonded silica-based materials suffer from two major limitations: first, residual silanol groups frequently have adverse effects on chromatographic performance, and second, silica-based materials are stable only over a pH range of 2-8.5. Polymeric supports exhibit enhanced pH stability but are often limited by their lack of structural rigidity and low efficiencies due to the poor diffusional properties of solutes in these materials.
As a result of the aforementioned difficulties, attention has been given to an approach that involves the deposition of a hydrophobic, chemically stable polymer onto the surface of an inorganic carrier followed by a radical-initiated cross-linking reaction which serves to immobilize a thin layer of polymer on the surface of the support. Methods for coating inorganic supports with polymers to create supports for chromatography are known. Schnecko and Bieber (Schnecko, H. and Bieber, O. Die Angewandte Mackromolekulare Chemie 1971, 20, 111-119) describe coating Chromosorb P with polymers including polybutadiene and hydroxy-terminated polybutadienes and nitrile rubber, as well as dibromo polybutadiene after amine treatment. The use of dicumyl peroxide is disclosed as an agent for subsequently crosslinking the polymers to create stationary phases for gas chromatography. This disclosure in inapplicable to the instant invention because of the differences between gas and liquid chromatography. For example, in modern high performance liquid chromatography (HPLC), pressures of several thousand lb/in.sup.2 are often developed with the chromatography columns. The high pressure liquid within the column subjects the base material to conditions which are much more severe, or substantially different from those experienced in gas chromatography. Unlike liquid chromatography, gas chromatography can not be applied to the separation of solids, such as proteins and peptides, because they can not be entrained in the mobile gas phase.
Schomburg et al. (Schomburg, G.; Kohler, J; Figge, H.; Deege, A.; Bien-Vogelsang, U., Chromatographia 1984, 18, 265-274) report the immobilization of polymers on particles of silica and alumina using .sup.60 Co irradiation to prepare stationary phases for liquid chromatography and describe further improvements in subsequent publications (including Bien-Vogelsang, U.; Deege, A.; Figge, H.; Khler, J.; Schomburg, G., Chromatographia 1984, 19, 170-179; Figge, H.; Deege, A.; Kohler, J.; Schomburg, G. J. Chromatography 1986, 351, 393-408; Kolla, P, Kohler, J. Schomburg, G. Chromatographia 1987, 23, 465-472).
Kosaka et al. (U.S. Pat. No. 4,054,353, Aug. 30, 1977) describe the radiation-induced crosslinking of monomers on the surface of inorganic substrates and simple chemical modification of some polymers, such as sulfonation of styrene. Berezkin et al. (Berezkin, V. G.; Kolbanovskii, Yu. A.; Kyazimov, E. A. Zh. Fiz. Khim. 1966, 40, 1921) also describe modifying supports by depositing monomers and crosslinking by irradiation.
Regnier et al. in a series of publications and in a patent (U.S. Pat. No. 4,245,005, Jan. 13, 1981) describe the adsorption of coatings, such as amines, to inorganic supports and then the crosslinking of the coatings by chemical means to create stationary phases suitable for ion-exchange chromatography.
A disadvantage of these methods is that they produce stationary phases which exhibit chromatographic behavior different from that of commonly-used, commercially-available materials. There is a great reluctance among those who practice chromatography to use stationary phases that exhibit unfamiliar behavior because the large existing body of knowledge developed based on the use of certain stationary phases is not applicable. Accordingly, it would be very desirable to have a stationary phase that exhibits chromatographic behavior similar to commonly-used, commercially-available materials, but which also exhibits improved chemical and mechanical stability to alleviate the deficiencies of inorganic supports coated with organosilanes and of polymeric supports. The materials described by this invention may be shown to exhibit excellent stability under acidic and basic conditions, high efficiency and good chromatographic selectivities especially for proteins and peptides.